We evaluated the antimicrobial activity of fosfomycin against a randomly selected sample of 30 Klebsiella pneumoniae, 30 Pseudomonas aeruginosa, and 30 Acinetobacter baumannii multidrug-resistant, clinical isolates from patients in a general tertiary care hospital in Athens, Greece. Standard laboratory methods were used for susceptibility testing to commonly used antibiotics and the detection of extended-spectrum-beta-lactamase (ESBL) and metallo-beta-lactamase (MBL) production. The minimum inhibitory concentration (MIC) of fosfomycin for each isolate was determined by the agar dilution method. All K. pneumoniae isolates were both ESBL and MBL producers; all P. aeruginosa isolates were ESBL producers. The K. pneumoniae strains had fosfomycin MICs distributed across a range of 8-64 microg/ml; MIC(50) was 16 microg/ml and MIC(90) 32 microg/ml. The fosfomycin MICs of the P. aeruginosa strains had a distribution across a range of 4 to over 512 microg/ml; MIC(50) was 32 microg/ml and MIC(90) 128 microg/ml. The fosfomycin MICs of the A. baumannii strains had a distribution across a range of 64 to over 512 microg/ml; MIC(50) was 256 microg/ml and MIC(90) more than 512 microg/ml. Although standardized fosfomycin MIC interpretative breakpoints for the species studied are lacking, the findings of our study support the idea that fosfomycin may be further investigated as one among a decreasing list of therapeutic options for the treatment of infections due to multidrug-resistant strains of, primarily, K. pneumoniae and, secondly, P. aeruginosa.